Tacrolimus is an immunosuppressive agent produced by Streptomyces tsukubaensis No. 9993 and is the compound of formula (I) wherein R.sub.1 and R.sub.2 are both hydrogen. Tacrolimus, which is also called FK-506, has first discovered by Tanaka, Kuroda and their colleague in Japan see, J. Am. Chem. Soc., 1987, 109, 5031 and U.S. Pat. No. 4,894,366 issued on Jan. 16, 1990!.
Tacrolimus and the related compounds have been shown to be useful in treating obstructive bronchial diseases, particularly asthma, male pattern alopecia, senile alopecia, rheumatoid arthritis, diabetic diseases, posterior uveitisophthalmic diseases, hepatic disorder associated with ischemia, allergic encephalomyelitis, glomerulonephritis, systemic erythematosus lupus, polypharmaceutic resistance, inflammation of mucosal membrane and blood vessel, cytomegaloviral infection and idiopathic thrombocytopenic purpura, hyperthyroidism, etc.
Tacrolimus is combined with intracellular protein FKBP-12 to form a complex of tacrolimus-FKBP-12, Ca, calmodulin and calcineurin, which inhibits phosphate activation of calcineurin and, as a result, inhibits the production of intranuclear factor of activated T cells (NF-AT), which is regarded as a substance initiating genetic transcription for lymphokine (interleukin-2, gamma-interferon) production, to inhibit the activation of T-lymphocytes. Cyclosporin and rapamycin also have been shown to be effective as immunosuppressive agents, and therefore, are useful in preventing transplantation rejection see, FASEB, 1989, 3, 3411; FASEB, 1989, 3, 5256; Calne, R. et al., Lancet, 1978, 1183; and U.S. Pat. No. 5,180,899!.
Tacrolimus is a neutral substance and is generally well dissolved in organic solvents but not in water and n-hexane. Due to such low solubility (12 .mu.g/ml) in water, in formulating the composition castor oil derivatives are required to dissolve tacrolimus. For example, in intravenous injections of tacrolimus 200 mg/ml of hydrogenated polyoxy 60 castor oil (HCO-60) and 80% (v/v) absolute alcohol are required as the solubilizing aid for dissolving 5 mg of tacrolimus.
Substantially no patient who received tacrolimus injection has experienced anaphylaxis. However, other pharmaceutical compositions containing castor oil derivatives have caused anaphylaxis in few patients. Therefore, tacrolimus injection is required for patients who cannot take tacrolimus capsules due to a potential risk of such anaphylaxis. In addition, the use of non-aqueous solvents such as ethanol, propylene glycol or polyethylene glycol 400 in parenteral preparations may cause side effects such as hemolysis, local irritation at injection site, etc., and needs to be carefully considered.
Polyethylene glycol (PEG) is a linear or branched, neutral polymer available in a variety of molecular weights and is well soluble in water and methylene chloride. At molecular weights less than 1000 PEGs are viscous, colorless liquids; and higher molecular weight PEGs are waxy, white solids. The melting point of the solid is proportional to the molecular weight. PEGs having molecular weights ranging from a few hundred to approximately 20,000 are commonly used in biological and biotechnological applications.
Of much interest in the biomedical areas is the fact that polyethylene glycol is nontoxic and was approved by FDA for internal consumption. Polyethylene glycol is very widely used in the field of pharmaceuticals, cosmetics and personal sanitary products. One of the most extensively studied drug delivery systems is the covalent binding of methoxy polyethylene glycol to the surface of protein.
Water soluble polymers such as polyethylene glycol (PEG) and methoxy polyethylene glycol (mPEG) are used for binding the non-aqueous drugs. Therefore, the present invention has developed the novel tacrolimus conjugated compound, which can be dissolved in water, formed by chemically binding the sparingly soluble drug, tacrolimus, with the water soluble polymer, methoxy polyethylene glycol. The conjugated compound of the present invention can be used as an immunosuppressive, antiinflammatory, antifungal, antiproliferative and antitumor agent.